Randomness and optimality in enhanced DNA ligation with crowding effects

TL;DR

This paper introduces a new statistical analysis method for DNA ligation, demonstrating that molecular crowding can suppress exponential decay and optimize ligation efficiency for synthesizing long DNA sequences.

Contribution

The study presents a novel qPCR-based statistical analysis of DNA ligation randomness and reveals how crowding effects can enhance long DNA synthesis by balancing ligation speed and enzyme availability.

Findings

Crowding suppresses exponential decay in DNA ligation.

Optimal crowding conditions activate ligation.

Molecular attraction under crowding improves long DNA synthesis.

Abstract

Enzymatic ligation is essential for the synthesis of long DNA. However, the number of ligated products exponentially decays as the DNA synthesis proceeds in a random manner. The controlling of ligation randomness is of importance to suppress exponential decay and demonstrate an efficient synthesis of long DNA. Here, we report the analysis of randomness in sequential DNA ligations, named qPCR-based STatistical Analysis of Randomness (qPCR-bSTAR), by a probability distribution of ligated DNA concentration. We show that the exponential decay is suppressed in a solution of another polymer and DNA ligation is activated at an optimal crowded condition. Theoretical model of kinetic ligation explains that intermolecular attraction due to molecular crowding can be involved in the optimal balance of the ligation speed and the available ligase. Our finding indicates that crowding effects enhance the synthesis of long DNA that retains large genetic information.